Fig. 4: Interaction of INSM1 with TEAD1.

A Western blot images for the Co-IP experiments between INSM1 and TEAD1 in HeLa and H69 cells. IP was carried out using anti-Flag antibody for Flag-fused TEAD1 in HeLa cell, and using anti-INSM1 antibody in H69 cell. The experiment was repeated independently for three times with similar results. B BiFC between cYFP-fused TEAD1 and different variants of INSM1 fused with nYFP in HeLa cell. Bar, 20 μm. The experiment was repeated independently for three times with similar results. C Y2H assay between different variants of INSM1 and TEAD1. Transformants harboring both AD- and BD-derived constructs were grown on SD2 (–Trp/–Leu) medium for growth control and SD4 (–Trp/–Leu/–His/–Ade) medium for the interaction test. D Binding curve and K_D value between INSM1-ZF1 and TEAD1-TEAD determined by FP assay. Source data are provided as a Source Data file. E The fitted AUC data of INSM1-ZF1, TEAD1-TEAD, and their complex in sedimentation velocity experiments. The determined molecular weight for each peak is shown as inset. Asterisk marks the unspecific peak. Source data are provided as a Source Data file. F Overlay of a series of ¹H−¹⁵N HSQC spectra of ¹⁵N-labeled INSM1-ZF1 in the presence of unlabeled TEAD1-TEAD at different molar ratios, which are colored differently as indicated. G Overlay of a series of ¹H-¹⁵N HSQC spectra of ¹⁵N-labeled TEAD1-TEAD in the presence of unlabeled INSM1-ZF1 at different molar ratios, which are colored differently as indicated. Chemical shift perturbation (CSP) for each residue of INSM1-ZF1 (H) and TEAD1-TEAD (I) during NMR titration is calculated and illustrated with the secondary structure elements shown on top. CSP value of 0.1 was selected as the significant level for globally comparison. Pink-filled columns mark the residues with disappeared signals during titration. The residues with CSP over 0.1 and those with disappeared signals were labelled in (F, G). Unassigned residues are marked with asterisks. P, proline. The residues with hardly any CSP are marked with cycles. Source data are provided as a Source Data file. The residues with CSP values over 0.1 are shown on the structures of INSM1-ZF1 (J, blue) and TEAD1-TEAD (K, orange) in cartoon and surface views, respectively. Residues with disappeared peaks in the spectra are shown and marked in pink.